A MUTATIONAL ANALYSIS OF RECEPTOR-BINDING SITES OF INTERLEUKIN-1-BETA- DIFFERENCES IN BINDING OF HUMAN INTERLEUKIN-1-BETA MUTEINS TO HUMANAND MOUSE RECEPTORS

The 3-D crystal structure of interleukin-1 beta (IL-1 beta) has been u sed to define its receptor binding surface by mutational analysis. The surface of IL-1 beta was probed by site-directed mutagenesis. A total of 27 different IL-1 beta muteins were constructed, purified and anal yzed. Receptor binding measurements on mouse and human cell lines were performed to identify receptor affinities. IL-1 beta muteins with mod ified receptor affinity were evaluated for structural integrity by CD spectroscopy or X-ray crystallography. Changes in six surface loops, a s well as in the C- and N-termini, yielded muteins with lower binding affinities. Two muteins with intact binding affinities showed 10- to 1 00-fold reduced biological activity. The surface region involved in re ceptor binding constitutes a discontinuous area of similar to 1000 Ang strom(2) formed by discontinuous polypeptide chain stretches. Based on these results, a subdivision into two distinct local areas is propose d. Differences in receptor binding affinities for human and mouse rece ptors have been observed for some muteins, but not for wild-type IL-1 beta. This is the first time a difference in binding affinity of IL-1 beta muteins to human and mouse receptors has been demonstrated.